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Increased gene dosage for β - and κ -casein in transgenic cattle improves milk composition through complex effects

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ABSTRACT

We have previously generated transgenic cattle with additional copies of bovine β- and κ casein genes. An initial characterisation of milk produced with a hormonally induced lactation from these transgenic cows showed an altered milk composition with elevated β-casein levels and twofold increased κ-casein content. Here we report the first in-depth characterisation of the composition of the enriched casein milk that was produced through a natural lactation. We have analyzed milk from the high expressing transgenic line TG3 for milk composition at early, peak, mid and late lactation. The introduction of additional β- and κ-casein genes resulted in the expected expression of the transgene derived proteins and an associated reduction in the size of the casein micelles. Expression of the transgenes was associated with complex changes in the expression levels of other milk proteins. Two other major milk components were affected, namely fat and micronutrients. In addition, the sialic acid content of the milk was increased. In contrast, the level of lactose remained unchanged. This novel milk with its substantially altered composition will provide insights into the regulatory processes synchronizing the synthesis and assembly of milk components, as well as production of potentially healthier milk with improved dairy processing characteristics.

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Two-dimensional electrophoresis of mammary secretion from a transgenic (TG3) and a control (WT) cow.Mammary secretion was collected from a TG3 and a wild type cow, firstly as one-year old heifers after hormonal induction of lactation (induced), and then from the same animals at peak lactation after a natural calving (natural). Milk proteins were separated by isoelectric point (pI) in the first dimension and molecular weight (MW) in the second dimension. The gels were stained for protein with Coomassie blue and sections of each gel are shown in which the major caseins (as marked) are resolved. The white centred arrow indicates the exogenously expressed A3 isoform of β-casein. The black centred arrow indicates the exogenously expressed B isoform of κ-casein. Shown are representative gels which have been cropped for improved clarity. The full size gels are presented in Supplementary Fig. S2.
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f1: Two-dimensional electrophoresis of mammary secretion from a transgenic (TG3) and a control (WT) cow.Mammary secretion was collected from a TG3 and a wild type cow, firstly as one-year old heifers after hormonal induction of lactation (induced), and then from the same animals at peak lactation after a natural calving (natural). Milk proteins were separated by isoelectric point (pI) in the first dimension and molecular weight (MW) in the second dimension. The gels were stained for protein with Coomassie blue and sections of each gel are shown in which the major caseins (as marked) are resolved. The white centred arrow indicates the exogenously expressed A3 isoform of β-casein. The black centred arrow indicates the exogenously expressed B isoform of κ-casein. Shown are representative gels which have been cropped for improved clarity. The full size gels are presented in Supplementary Fig. S2.

Mentions: We have previously reported the generation of the transgenic cattle line TG3, harbouring additional genes for bovine β- and κ-casein and described their expression potential deduced from a hormonally induced lactation16. To confirm that the transgene derived milk proteins β-casein A3 and κ-casein B are expressed in natural lactation milk at similar levels to what we have detected in hormonally induced milk we initially analyzed early lactation milk samples from the first natural lactation of seven TG3 founder cows and three wild type control cows with the same genetic background. Two-dimensional (2D) analysis of milk proteins revealed a similar pattern of relative abundance for induced and natural milk (Fig. 1). The transgene derived β-casein A3 and κ-casein B variants were enhanced in abundance in natural transgenic milk compared to induced transgenic milk. In contrast to induced TG3 milk, in natural TG3 milk the transgene-derived caseins were the most highly expressed variants of β-casein and κ-casein, well above the expression levels of the endogenous variants β-casein A1 and A2 and κ-casein A. In addition, the relatively high abundance of κ-casein B resulted in a more pronounced representation of the glycosylated κ-casein isoforms in the natural milk sample from the TG3 cows compared to the induced milk.


Increased gene dosage for β - and κ -casein in transgenic cattle improves milk composition through complex effects
Two-dimensional electrophoresis of mammary secretion from a transgenic (TG3) and a control (WT) cow.Mammary secretion was collected from a TG3 and a wild type cow, firstly as one-year old heifers after hormonal induction of lactation (induced), and then from the same animals at peak lactation after a natural calving (natural). Milk proteins were separated by isoelectric point (pI) in the first dimension and molecular weight (MW) in the second dimension. The gels were stained for protein with Coomassie blue and sections of each gel are shown in which the major caseins (as marked) are resolved. The white centred arrow indicates the exogenously expressed A3 isoform of β-casein. The black centred arrow indicates the exogenously expressed B isoform of κ-casein. Shown are representative gels which have been cropped for improved clarity. The full size gels are presented in Supplementary Fig. S2.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5120311&req=5

f1: Two-dimensional electrophoresis of mammary secretion from a transgenic (TG3) and a control (WT) cow.Mammary secretion was collected from a TG3 and a wild type cow, firstly as one-year old heifers after hormonal induction of lactation (induced), and then from the same animals at peak lactation after a natural calving (natural). Milk proteins were separated by isoelectric point (pI) in the first dimension and molecular weight (MW) in the second dimension. The gels were stained for protein with Coomassie blue and sections of each gel are shown in which the major caseins (as marked) are resolved. The white centred arrow indicates the exogenously expressed A3 isoform of β-casein. The black centred arrow indicates the exogenously expressed B isoform of κ-casein. Shown are representative gels which have been cropped for improved clarity. The full size gels are presented in Supplementary Fig. S2.
Mentions: We have previously reported the generation of the transgenic cattle line TG3, harbouring additional genes for bovine β- and κ-casein and described their expression potential deduced from a hormonally induced lactation16. To confirm that the transgene derived milk proteins β-casein A3 and κ-casein B are expressed in natural lactation milk at similar levels to what we have detected in hormonally induced milk we initially analyzed early lactation milk samples from the first natural lactation of seven TG3 founder cows and three wild type control cows with the same genetic background. Two-dimensional (2D) analysis of milk proteins revealed a similar pattern of relative abundance for induced and natural milk (Fig. 1). The transgene derived β-casein A3 and κ-casein B variants were enhanced in abundance in natural transgenic milk compared to induced transgenic milk. In contrast to induced TG3 milk, in natural TG3 milk the transgene-derived caseins were the most highly expressed variants of β-casein and κ-casein, well above the expression levels of the endogenous variants β-casein A1 and A2 and κ-casein A. In addition, the relatively high abundance of κ-casein B resulted in a more pronounced representation of the glycosylated κ-casein isoforms in the natural milk sample from the TG3 cows compared to the induced milk.

View Article: PubMed Central - PubMed

ABSTRACT

We have previously generated transgenic cattle with additional copies of bovine β- and κ casein genes. An initial characterisation of milk produced with a hormonally induced lactation from these transgenic cows showed an altered milk composition with elevated β-casein levels and twofold increased κ-casein content. Here we report the first in-depth characterisation of the composition of the enriched casein milk that was produced through a natural lactation. We have analyzed milk from the high expressing transgenic line TG3 for milk composition at early, peak, mid and late lactation. The introduction of additional β- and κ-casein genes resulted in the expected expression of the transgene derived proteins and an associated reduction in the size of the casein micelles. Expression of the transgenes was associated with complex changes in the expression levels of other milk proteins. Two other major milk components were affected, namely fat and micronutrients. In addition, the sialic acid content of the milk was increased. In contrast, the level of lactose remained unchanged. This novel milk with its substantially altered composition will provide insights into the regulatory processes synchronizing the synthesis and assembly of milk components, as well as production of potentially healthier milk with improved dairy processing characteristics.

No MeSH data available.


Related in: MedlinePlus